Arsenic trioxide (ATO) is a Food and Drug Administration (FDA) approved therapy for a subtype of acute myeloid leukemia, acute promyelocytic leukemia (APL). There is also potential for clinical development of arsenic and/or other medicinal compounds for the treatment of other malignancies. However, to date arsenic has not shown significant clinical activity as a single agent outside of APL, likely due to the activation of negative feedback pathways that counteract its activity. The major goal of our research group is to investigate the mechanisms of action of arsenic in myeloid leukemias, understand the negative feedback pathways activate by arsenic and identify agents that can be combined with it to enhance clinical outcomes in patients. In this proposal we seek to investigate the role of AMPK in the anti-leukemic effects of arsenic and how that contributes to the induction of mTOR as a negative feedback loop as well as in the induction of autophagy. It was previously shown in our laboratory that during treatment of leukemia cells with arsenic there is activation of the mTOR pathway through an unknown mechanism. We have preliminary data to suggest that arsenic directly binds to AMPK and inhibits its activity. Because AMPK is a direct negative regulator of mTOR, we propose that this is an important mechanism for activation of the mTOR pathway by arsenic. In Aim 1 we will investigate how arsenic affects AMPK structure and function. We also propose that arsenic through AMPK inhibition leads to induction of autophagy through the VPS34/Beclin/UVRAG complex since AMPK can negative regulate VSP34. In Aim 2 we will combine arsenic with AMPK activators to investigate the role of AMPK signaling in the induction of anti-leukemic effects of arsenic in vivo in a xenograft mouse model of acute myeloid leukemia (AML). Altogether, this work should define the mechanisms by which arsenic activates negative feedback pathways and may lead to the development of novel agents for the treatment of leukemias.